scholarly journals Detection of uncoupled circadian rhythms in individual cells of Lemna minor using a dual-color bioluminescence monitoring system

2021 ◽  
Author(s):  
Emiri Watanabe ◽  
Minako Isoda ◽  
Tomoaki Muranaka ◽  
Shogo Ito ◽  
Tokitaka Oyama
Keyword(s):  
Circadian Rhythms ◽  
Monitoring System ◽  
Lemna Minor ◽  
Large Cell ◽  
Firefly Luciferase ◽  
Circadian Oscillation ◽  
Mutant Form ◽  
Oscillation System ◽  
Pass Filter ◽  
Dual Color

Abstract The plant circadian oscillation system is based on the circadian clock of individual cells. Circadian behavior of cells has been observed by monitoring the circadian reporter activity such as bioluminescence of AtCCA1::LUC+. To deeply analyze different circadian behaviors in individual cells, we developed the dual-color bioluminescence monitoring system that automatically measured the luminescence of two luciferase reporters simultaneously at a single-cell level. We selected a yellow-green-emitting firefly luciferase (LUC+) and a red-emitting luciferase (PtRLUC) that is a mutant form of Brazilian click beetle ELUC. We used AtCCA1::LUC+ and CaMV35S::PtRLUC. CaMV35S::LUC+ was previously reported as a circadian reporter with a low amplitude rhythm. These bioluminescent reporters were introduced into the cells of a duckweed, Lemna minor, by particle bombardment. Time series of the bioluminescence of individual cells in a frond were obtained using a dual-color bioluminescence monitoring system with a green-pass- and red-pass filter. Luminescence intensities from the LUC+ and PtRLUC of each cell were calculated from the filtered luminescence intensities. We succeeded in reconstructing the bioluminescence behaviors of AtCCA1::LUC+ and CaMV35S::PtRLUC in the same cells. Under prolonged constant light conditions, AtCCA1::LUC+ showed a robust circadian rhythm in individual cells in an asynchronous state in the frond, as previously reported. In contrast, CaMV35S::PtRLUC stochastically showed circadian rhythms in a synchronous state. These results strongly suggested the uncoupling of cellular behavior between these circadian reporters. This dual-color bioluminescence monitoring system is a powerful tool to analyze various stochastic phenomena accompanying large cell-to-cell variation in gene expression.

scholarly journals Detection of uncoupled circadian rhythms in individual cells of Lemna minor using a dual-color bioluminescence monitoring system

2020 ◽  
Author(s):  
Emiri Watanabe ◽  
Minako Isoda ◽  
Tomoaki Muranaka ◽  
Shogo Ito ◽  
Tokitaka Oyama
Keyword(s):  
Circadian Rhythms ◽  
Single Cell ◽  
Monitoring System ◽  
Lemna Minor ◽  
Single Cell Level ◽  
Cell Level ◽  
Dual Color ◽  
Stochastic Phenomena ◽  
Circadian Behavior ◽  
Bioluminescent Reporters

SummaryThe plant circadian oscillation system is based on the circadian clock of individual cells and coordinates the circadian behavior of the plant body. To observe the cellular circadian behavior of both the oscillator and its output in plants, we developed the dual-color bioluminescence monitoring system that automatically measured the luminescence of two luciferase reporters simultaneously at a single-cell level. We selected a yellow-green-emitting firefly luciferase (LUC+) and a red-emitting luciferase (PtRLUC) that is a mutant form of Brazilian click beetle ELUC. We used AtCCA1::LUC+ and CaMV35S::PtRLUC to observe the cellular behavior of the oscillator and output, respectively. These bioluminescent reporters were introduced into the cells of a duckweed, Lemna minor, by particle bombardment. Time series of the bioluminescence of individual cells in a frond were obtained using a dual-color bioluminescence monitoring system with a green-pass- and red-pass filter. Luminescence intensities from the LUC+ and PtRLUC of each cell were calculated from the filtered luminescence intensities. We succeeded in reconstructing the bioluminescence behaviors of AtCCA1::LUC+ and CaMV35S::PtRLUC in the same cells. Under prolonged constant light conditions, AtCCA1::LUC+ showed a robust circadian rhythm in individual cells in an asynchronous state in the frond, as previously reported in studies using other plants. In contrast, CaMV35S::PtRLUC stochastically showed circadian rhythms in a synchronous state. Thus, we clearly demonstrated the uncoupling between the oscillator and output in individual cells. This dual-color bioluminescence monitoring system is a powerful tool to analyze various stochastic phenomena accompanying large cell-to-cell variation in gene expression.Significance statementWe succeeded in establishing the world’s first dual-color bioluminescence monitoring system at a single-cell level that enables simultaneous measurement of the luminescence activities of two reporter genes in plants. This system is a strong tool to analyze stochastic phenomena, and we clearly demonstrated the uncoupling of rhythmic behavior between two bioluminescent reporters in individual cells that stochastically occurred in the same plant.

Design of Fiber Distributed Busway Temperature Monitoring System

2013 ◽  
Vol 694-697 ◽  
pp. 1114-1117
Author(s):  
Yun Qin
Keyword(s):  
Monitoring System ◽  
Temperature Monitoring ◽  
Pass Filter ◽  
Low Pass Filter ◽  
Arithmetic Average ◽  
Low Snr ◽  
Current State ◽  
Low Pass ◽  
Temperature Measurement System ◽  
Scattering Signal

The reliability of busway in high current state is closely related to its temperature. So effectively monitoring temperature is the key issue to ensure the busway work safely. Because of the constraints such as high voltage and strong electromagnetic fields, traditional temperature measurement system can not meet the requirements. A distributed busway temperature monitoring system is designed in this paper based on the temperature effect of fiber Raman scattering signal. Arithmetic average, wavelet noise reduction and low-pass filter combination of data processing is used to overcome the very low SNR. Experiments show that the system can measure the temperature of 1km busway effectively. Temperature resolution can achieve 3°C.

scholarly journals Time-of-day-dependent responses of cyanobacterial cellular viability against oxidative stress

2019 ◽  
Author(s):  
Kenya Tanaka ◽  
Ginga Shimakawa ◽  
Shuji Nakanishi
Keyword(s):  
Oxidative Stress ◽  
Circadian Rhythms ◽  
Cell Viability ◽  
Circadian Clock ◽  
Stress Tolerance ◽  
Time Of Day ◽  
Circadian Oscillation ◽  
Subjective Time ◽  
Rhythmic Pattern ◽  
Oxidative Stress Tolerance

AbstractAs an adaptation to periodic fluctuations of environmental light, photosynthetic organisms have evolved a circadian clock. Control by the circadian clock of many cellular physiological functions, including antioxidant enzymes, metabolism and the cell cycle, has attracted attention in the context of oxidative stress tolerance. However, since each physiological function works in an integrated manner to deal with oxidative stress, whether or not cell responses to oxidative stress are under circadian control remains an open question. In fact, circadian rhythms of oxidative stress tolerance have not yet been experimentally demonstrated. In the present work, we applied an assay using methyl viologen (MV), which generates reactive oxygen species (ROS) under light irradiation, and experimentally verified the circadian rhythms of oxidative stress tolerance in photosynthetic cells of the cyanobacterium Synechococcus elongatus PCC7942, a standard model species for investigation of the circadian clock. Here, we report that ROS generated by MV treatment causes damage to stroma components and not to the photosynthetic electron transportation chain, leading to reduced cell viability. The degree of decrease in cell viability was dependent on the subjective time at which oxidative stress was applied. Thus, oxidative stress tolerance was shown to exhibit circadian rhythms. In addition, the rhythmic pattern of oxidative stress tolerance disappeared in mutant cells lacking the essential clock genes. Notably, ROS levels changed periodically, independent of the MV treatment. Thus, we demonstrate for the first time that in cyanobacterial cells, oxidative stress tolerance shows circadian oscillation.

scholarly journals Sensitive Dual Color In Vivo Bioluminescence Imaging Using a New Red Codon Optimized Firefly Luciferase and a Green Click Beetle Luciferase

PLoS ONE ◽  
2011 ◽  
Vol 6 (4) ◽  
pp. e19277 ◽  
Author(s):  
Laura Mezzanotte ◽  
Ivo Que ◽  
Eric Kaijzel ◽  
Bruce Branchini ◽  
Aldo Roda ◽  
...  
Keyword(s):  
Bioluminescence Imaging ◽  
Firefly Luciferase ◽  
Dual Color ◽  
Click Beetle ◽  
In Vivo Bioluminescence Imaging

scholarly journals Circadian variations in gene expression in rat abdominal adipose tissue and relationship to physiology

2010 ◽  
Vol 42A (2) ◽  
pp. 141-152 ◽  
Author(s):  
Siddharth Sukumaran ◽  
Bai Xue ◽  
William J. Jusko ◽  
Debra C. DuBois ◽  
Richard R. Almon
Keyword(s):  
Gene Expression ◽  
Adipose Tissue ◽  
Circadian Rhythms ◽  
Regulation Of Gene Expression ◽  
Gene Array ◽  
Circadian Oscillation ◽  
Peripheral Tissues ◽  
Physiological Processes ◽  
Expression Of Genes ◽  
Rat Adipose Tissue

Circadian rhythms occur in all levels of organization from expression of genes to complex physiological processes. Although much is known about the mechanism of the central clock in the suprachiasmatic nucleus, the regulation of clocks present in peripheral tissues as well as the genes regulated by those clocks is still unclear. In this study, the circadian regulation of gene expression was examined in rat adipose tissue. A rich time series involving 54 animals euthanized at 18 time points within the 24-h cycle (12:12 h light-dark) was performed. mRNA expression was examined with Affymetrix gene array chips and quantitative real-time PCR, along with selected physiological measurements. Transcription factors involved in the regulation of central rhythms were examined, and 13 showed circadian oscillations. Mining of microarray data identified 190 probe sets that showed robust circadian oscillations. Circadian regulated probe sets were further parsed into seven distinct temporal clusters, with >70% of the genes showing maximum expression during the active/dark period. These genes were grouped into eight functional categories, which were examined within the context of their temporal expression. Circadian oscillations were also observed in plasma leptin, corticosterone, insulin, glucose, triglycerides, free fatty acids, and LDL cholesterol. Circadian oscillation in these physiological measurements along with the functional categorization of these genes suggests an important role for circadian rhythms in controlling various functions in white adipose tissue including adipogenesis, energy metabolism, and immune regulation.

scholarly journals An endogenous basis for synchronization manners of the circadian rhythm in proliferating Lemna minor plants

2021 ◽  
Author(s):  
Kenya Ueno ◽  
Shogo Ito ◽  
Tokitaka Oyama
Keyword(s):  
Circadian Rhythm ◽  
Circadian Rhythms ◽  
Lemna Minor ◽  
Spatiotemporal Analysis ◽  
Cell Coupling ◽  
Long Distance ◽  
Detached Leaves ◽  
Circadian Organization ◽  
A Cell ◽  
Local Coupling

AbstractEndogenous circadian rhythms in plants play a role in adaptation to day-night cycles. The circadian clock is a cell-autonomous system that functions through the coordination of time information in the plant body. Synchronization of cellular clocks is based on coordination mechanisms; the synchronization manners in proliferating plants remain unclear. We performed spatiotemporal analysis of the circadian rhythm of fronds (leaf-like plant units) of proliferating Lemna minor plants carrying a circadian bioluminescence reporter, AtCCA1:LUC. Noninvasive observations of the bioluminescence of fast-growing two-dimensional plants allowed us to analyze the circadian rhythms at a cell-level resolution and obtain information regarding frond lineage. We focused on spontaneous circadian organization under constant light conditions for plants with light/dark treatment (LD-grown) or without it (LL-grown). Even fronds developing from an LL-grown parental frond showed coherent circadian rhythms among them. This allowed the maintenance of circadian rhythmicity in proliferating plants. Inside a frond, a centrifugal phase/period pattern was observed in LD-grown plants, whereas various phase patterns with traveling waves were formed in LL-grown plants. These patterns were model-simulated by local coupling of cellular circadian oscillators with different initial synchronous states in fronds. Taken together with similar patterning previously reported for detached leaves of Arabidopsis, it is strongly suggested that local coupling is the primary force for the development of these phase patterns in plants lacking long-distance communication. We propose a basic framework of spontaneous phase patterning with three stages of circadian organization: initial phasing, evolution of patterning, and desynchronization/randomizing of phase, in association with altering cell-cell coupling.

scholarly journals Purine Catabolism Shows a Dampened Circadian Rhythmicity in a High-fat Diet-Induced Mouse Model of Obesity

Molecules ◽  
2019 ◽  
Vol 24 (24) ◽  
pp. 4524 ◽  
Author(s):  
Runbin Sun ◽  
Jingqiu Huang ◽  
Na Yang ◽  
Jun He ◽  
Xiaoyi Yu ◽  
...  
Keyword(s):  
Circadian Rhythms ◽  
Mouse Model ◽  
Nutritional Status ◽  
Metabolic Pathways ◽  
High Fat Diet ◽  
Clock Genes ◽  
Circadian Rhythmicity ◽  
Circadian Oscillation ◽  
High Fat ◽  
Purine Catabolism

High-calorie diet, circadian rhythms and metabolic features are intimately linked. However, the mediator(s) between nutritional status, circadian rhythms and metabolism remain largely unknown. This article aims to clarify the key metabolic pathways bridging nutritional status and circadian rhythms based on a combination of metabolomics and molecular biological techniques. A mouse model of high-fat diet-induced obesity was established and serum samples were collected in obese and normal mice at different zeitgeber times. Gas chromatography/mass spectrometry, multivariate/univariate data analyses and metabolic pathway analysis were used to reveal changes in metabolism. Metabolites involved in the metabolism of purines, carbohydrates, fatty acids and amino acids were markedly perturbed in accordance with circadian related variations, among which purine catabolism showed a typical oscillation. What’s more, the rhythmicity of purine catabolism dampened in the high-fat diet group. The expressions of clock genes and metabolic enzymes in the liver were measured. The mRNA expression of Xanthine oxidase (Xor) was highly correlated with the rhythmicity of Clock, Rev-erbα and Bmal1, as well as the metabolites involved in purine catabolism. These data showed that a high-fat diet altered the circadian rhythm of metabolic pathways, especially purine catabolism. It had an obvious circadian oscillation and a high-fat diet dampened its circadian rhythmicity. It was suggested that circadian rhythmicity of purine catabolism is related to circadian oscillations of expression of Xor, Uox and corresponding clock genes.

scholarly journals Circadian rhythms in Per1, PER2 and Ca2+ of a solitary SCN neuron cultured on a microisland

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Yoshihiro Hirata ◽  
Ryosuke Enoki ◽  
Kaori Kuribayashi-Shigetomi ◽  
Yoshiaki Oda ◽  
Sato Honma ◽  
...  
Keyword(s):  
Circadian Rhythms ◽  
Glial Cell ◽  
Glial Cells ◽  
Large Fraction ◽  
Circadian Rhythmicity ◽  
Circadian Oscillation ◽  
Culture Dish ◽  
Intracellular Ca ◽  
Oscillatory Coupling ◽  
Dispersed Cells

AbstractCircadian rhythms in Per1, PER2 expression and intracellular Ca2+ were measured from a solitary SCN neuron or glial cell which was physically isolated from other cells. Dispersed cells were cultured on a platform of microisland (100–200 μm in diameter) in a culture dish. Significant circadian rhythms were detected in 57.1% for Per1 and 70.0% for PER2 expression. When two neurons were located on the same island, the circadian rhythms showed desynchronization, indicating a lack of oscillatory coupling. Circadian rhythms were also detected in intracellular Ca2+ of solitary SCN neurons. The ratio of circadian positive neurons was significantly larger without co-habitant of glial cells (84.4%) than with it (25.0%). A relatively large fraction of SCN neurons generates the intrinsic circadian oscillation without neural or humoral networks. In addition, glial cells seem to interrupt the expression of the circadian rhythmicity of intracellular Ca2+ under these conditions.

Relay Feedback Oscillator Design for Modeling Circadian Rhythms in Cyanobacteria

2011 ◽  
Author(s):  
Jaromir Fiser ◽  
Pavel Zitek ◽  
Jan Cerveny
Keyword(s):  
Circadian Rhythms ◽  
Negative Feedback ◽  
Stability Margin ◽  
State Variables ◽  
Relay Feedback ◽  
Pass Filter ◽  
Low Pass Filter ◽  
Single Loop ◽  
Low Pass ◽  
Autoregulatory Mechanism

The paper introduces a relay feedback oscillator for modeling circadian rhythms in cyanobacteria. The relay feedback oscillator is equipped with low pass filter F(jω), hysteresis-type relay and negative feedback. This negative feedback represents an autoregulatory mechanism of the circadian clock and the notion of this autoregulatory mechanism is based on the well-known Goodwin biochemical oscillator [1]. The relay is responsible for the mediation of both the activation and degradation of oscillator state variables (protein concentrations) and in this way the pacemaker is constituted. Later on, low pass filter poles are identified for the purpose of modeling auto-oscillations with the free running period of 24h and the method of the pole identification consists in an ultimate frequency test providing stability margin of a single-loop composed of the filter and the relay in the feedback. Next, a relay output / input ratio of amplitudes and hysteresis are found out by the graphical test of the single-loop on the stability margin which is carried out in Bode graph. Finally, the output correspondence of relay feedback oscillator model with Miyoshi oscillator [2] is provided because the Miyoshi oscillator is well recognized among biochemical oscillators for species of cyanobacteria.

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